Enzymes

Enzymes: Important Terminology I. Metabolism In order for cells to maintain homeostasis, they must constantly convert chemicals from one form to another, in order to produce necessary molecules, obtain usable molecules from food, and produce energy rich molecules These constantly occurring chemical reactions are collectively known as metabolism

Metabolism a term to collectively describe all the chemical reactions occurring constantly in the cell that maintain homeostasis in a cell or organism D. Metabolic Pathways are the orderly step-wise series of chemical reactions from the initial reactants to the final products E. One reaction leads to the next; highly structured reaction

Ex. photosynthesis, cellular respiration G. It is controlled by enzymes 1. Each step (i.e. each chemical reaction) within the metabolic pathway requires a specific enzyme   Step 1 Step 2 Step 3 Step 4 Step 5 Step 6 A B C D E F G Reactant Enz. 1 Enz. 2 Enz. 3 Enz. 4 Enz. 5 Enz. 6 Product

H. There are reasons why metabolic pathways exist: It is not possible in biological systems to have a single reaction that could produce complex molecules from simple reactants ex. 6CO2 + 6H2O  C6H12O6 + 6O2 would never happen in a cell in one step (photosynthesis requires many intermediate steps)

2. One pathway can lead to several Others intermediate products of one pathway can be starting reactant for another pathway b. Ex. A  B C D 3. Having more than one step means that there are more places where the overall reaction can be controlled    Y  Z AA BB   X A B C D E  L    H Q M I R O N P K J S

Enzyme = Biological Catalysts A protein that can speed up a chemical reaction without being consumed that speeds up a chemical reaction Enzymes are the sites of chemical reactions, but aren’t used up in the reaction or permanently changed by the reaction E.g. They can hold reactant molecules together long enough for them to react

Enzymes are highly specific (each enzyme speeds up only one reaction) Specificity arises from the protein’s complex three-dimensional structure No cellular reaction will occur without its specific enzyme to act as catalyst Enzymes have a groove/dimple into which only specific reactant molecules may enter called the ACTIVE SITE

Enzyme names usually end with the suffix “ase” or sometimes “sin” Named after the substrate on which the enzyme works on Ex. RNA polymerase - constructs RNA Ex. Lactase - breaks down lactose Ex. Trypsin – breaks down proteins 4. Ex. Pepsin – breaks down proteins

Substrate is the reactant in an enzyme’s reaction The equation for an enzyme-catalyzed reaction is always: Enzyme + Substrate  Enzyme Substrate Complex Enzyme + Product

What are Enzymes Made of? A protein part called an APOENZYME that gives it its specificity A cofactor which is a non-protein molecule or ion needed for proper enzyme function (“helpers”)

C. Coenzymes are organic cofactors 1. Bind to the enzyme and usually participate directly in the reaction 2. Are essential because they are part of a coenzyme’s structure

Serve as carriers for groups or electrons May participate in reaction by accepting or giving atoms to the reaction Ex. NAD (nicotinamide adenine dinucleotide) is a coenzyme of many oxidation-reduction reactions 4. Interact with the substrate molecule a. Ex. Weaken bonds in the substrate

Many water-soluble vitamins help make up parts of coenzymes Ex. Niacin (nicotinic acid) makes up part of coeznyme NAD+ b. Ex. Riboflavin (vitamin B2) makes up part of coeznyme FAD  

Models: Enzyme Function I. Lock and Key Model ANIMATION A. The enzymes and substrate fit perfectly Together B. Only specific substrate(s) will fit into the active site, and enzymes will only act on their substrate

The Induced-Fit Model Induced Fit Model is a refinement of the Lock and Key model The act of binding the substrate(s) induces slight changes in shape that accommodates the substrate more perfectly and facilitates the chemical reaction about to take place

Upon binding, the enzyme undergoes a slight conformational change to more perfectly bind the substrates. D. Then the reaction takes place, the enzyme-substrate complex separates, and the enzyme reassumes its original shape

E. Lock and Key model vs. Induced Fit model ex. Lock and Key b. ex. Induced Fit

How does an Enzyme Work? Enzymes lowers the ACTIVATION ENERGY required for the reaction to proceed

Activation Energy is defined as the energy that must be supplied to cause molecules to react with one another Enzymes do this by bringing the substrate molecules together and holding them long enough for the reaction to take place D. E.g. Reactions that occur at 100C can occur at 37C with the use of an enzyme

Factors Affecting Enzyme Activity Enzymes are Proteins Enzyme are affected by the same things that affect proteins B. Since the shape of enzymes determines the shape of the active site, which determines their function, anything that changes the shape of an enzyme with affect the enzymatic yield.

C. Some factors are: 1. pH 2. Temperature 3.Concentrations of substrates 4.Concentration of enzyme 5.Presence of inhibitors

II. pH A. Enzyme shape is dependent upon pH- sensitive interactions between R-groups

B. Any change in pH denatures the enzyme by causing it to change shape: A denatured protein is one that has lost its normal configuration, and therefore its ability to form an enzyme-substrate complex 2. Most enzymes prefer pH’s of 6 – 8 3. Some exceptions: a. Pepsin in the stomach - pH ~ 2 b. Trypsin in the small intestine - pH ~ 8  

III. Temperature Temperature is a measure of the average kinetic energy in a container of molecules As the temperature rises, the reaction rates increase

Increasing the temperature slightly will, at first, increase the rate of reaction and product formation because it speeds up the rate at which substrates bump into enzymes 37C is optimum for human enzymes

Temperature too high (above about 40 °C) will denature the enzyme ANIMATION Slight denaturation causes the enzyme reactions to slow 2. Huge denaturation causes the enzyme reactions to stop

As the temperature drops, the reaction rates decreases because there are fewer enzyme-substrate collisions H. Very low temperatures do not normally denature the enzyme

IV. Concentrations of Substrates If the concentration of substrate increases, the amount of product increases However, after a certain concentration, the rate will not increase anymore, as all the enzymes are “saturated” with substrates and cannot work any faster

If the concentration of substrate decreases, the rate of product formation will generally decrease as well

Concentration of Enzyme This is what limits the overall rate of reaction. Providing there is adequate substrate (and their is typically millions more substrate molecules than enzyme molecules), the more enzyme you add, the more product you get

The less enzyme you have, the less product you get The less enzyme you have, the less product you get. In other words, if [enzyme] increases, rate of product formation increases. If the amount of enzyme decreases, the rate of product formation decreases. The rate will only level off if you run out of substrate, which is usually not the case.

Presence of Inhibitors Inhibitors are molecules that bind to the enzyme in some way to prevent or reduce the rate of substrate binding to enzyme

There are several ways in which inhibition can work: Competitive Inhibition: A molecule that looks like the substrate can compete for space at the active site (the place where the substrate binds to enzyme). This will slow down the reaction rate. The inhibitor binding to the Enzyme can be reversible or irreversible.

Obviously, the more inhibitors are added, the lower the rate of reaction, and the less product is going to be made.

Non-competitive Inhibition In this case, the inhibitor binds to another place on enzyme (not the active site). The inhibitor may look completely different from the substrate.

When the inhibitor binds, it causes the enzyme to change shape at the active site so substrate cannot bind. Binding may, as it is for competitive inhibition, be reversible or non-reversible. This type of inhibition is also known as “allosteric” inhibition.

Feedback Inhibition When an excess of product competitively binds with enzyme active sites, getting in the way of substrate molecules and slowing enzyme activity. This is called feedback inhibition. In a multi-enzyme pathway, often the end product binds at a special site on the first enzyme to shut down the whole pathway if the product builds up. This is Non-competitive inhibition or Allosteric control ANIMATION

Inhibitors can also be chemicals introduced into a system from the outside, and can act as medicines or poisons. Penicillin is a medicine that kills bacteria. It works by binding irreversibly to the enzyme that makes bacterial cell walls. HCN (hydrogen cyanide) is a lethal irreversible inhibitor of enzyme action in human. Lead (Pb++) and other heavy metals (like mercury (Hg++) and cadmium) are non-competitive inhibitors that cause poisoning when they bind irreversibly to enzymes and make them denature.

Cellular Respiration both plants and animals respire cells obtain their energy from the breakdown of carbohydrates, proteins and lipids these molecules are converted to glucose which is then broken down into CO2 and H2O C6H12O6 + 6O2  6CO2 + 6H2O

process of respiration is not as simple as indicated by the above reaction ANIMATION it requires 4 subpathways before CO2 and H2O are given off

These subpathways are: Glycolysis :the breakdown of glucose (6C) into 2 pyruvic acid molecules (2 - 3C) occurs in the cytoplasm end of the glycolysis process yields: 1. 2 pyruvic acid (3C) molecules 2. 4 ATP (a net of 2 ATP because 2 ATP was used to start the reaction) 2 NADH per glucose ANIMATION

2C segment being passed to the Krebs 1 NADH molecule per pyruvic acid 2. Transition reaction pyruvic acid (3C) is changed to active acetate (2C) (same as acetyl CoA) loss of 1C per molecule of pyruvic acid occurs in mitochondrion end of the transition reaction yields: 3. 1 CO2 2C segment being passed to the Krebs 1 NADH molecule per pyruvic acid 1 CO2

oxidative decarboxylation takes place (H's and C's removed) Krebs cycle ANIMATION oxidative decarboxylation takes place (H's and C's removed) cyclic; occurs over and over The end of Kreb Cycle yields: 1. 1 Oxaloacetic Acid molecule (4C) 2. 1 ATP 3. 3 NADH 4. 1 FADH2 5. 2 CO2

4. Respiratory chain ANIMATION H's (attached to NAD and FAD) are attached to O's to form H2O end of Electron Transport Phosphorylation yields: 1. 3 ATP per NADH 2. 2 ATP per FADH2 3. 2 water molecules per NADH or FADH2

energy produced from ONE glucose molecule Summary energy produced from ONE glucose molecule 1. NADH Subpathway Per molecule Total Glycolysis 1 NADH/ Glyceraldehyde 3- Phosphate 2 NADH   Transition Reaction 1 NADH/ pyruvic acid Kreb Cycle 3 NADH/ 6 NADH 10 NADH

2. FADH Subpathway Per molecule Total Kreb Cycle 1 FADH2/ pyruvic acid 2 FADH2   3. ATP Subpathway Per molecule Total Glycolysis 2 ATP/ Glyceraldehyde 3- Phosphate net of 2 ATP *used 2 ATP to start the reaction Kreb Cycle 1 ATP/cycle 2 ATP Electron Transport 3 ATP/NADH 30 ATP 2 ATP/FADH 4 ATP   38 ATP

Other Types of Metabolism carbohydrates (ie. glucose) are not the only products that can supply energy to the cell for storage as ATP proteins and lipids can also be broken down and supply energy to the cell fats are a much better supplier of energy than is glucose but glucose is the preferred energy source

Aerobic vs. Anerobic Respiration when oxygen is present (aerobic conditions), most organisms will undergo the Kreb's Cycle and Electron Transport Phosphorylation to produce ATP in eukaryotes, these processes occur in the mitochondria, while in prokaryotes they occur in the cytoplasm

under anaerobic conditions, the absence of oxygen, pyruvic acid can be routed by the organism into one of three pathways lactic acid fermentation alcohol fermentation 3.cellular (anaerobic) respiration

Humans cannot ferment alcohol in their own bodies, we lack the genetic information to do so. However, this is possible in something like yeast or anaerobic bacteria: these biochemical pathways, with their myriad reactions catalyzed by reaction-specific enzymes all under genetic control, are extremely complex.

alcohol fermentation is the formation of alcohol from sugar many organisms will also ferment pyruvic acid into other chemicals, such as lactic acid ex. Humans ferment lactic acid in muscles where oxygen becomes depleted, resulting in localized anaerobic conditions this lactic acid causes the muscle stiffness couch-potatoes feel after beginning exercise programs

the stiffness goes away after a few days since the cessation of strenuous activity allows aerobic conditions to return to the muscle, and the lactic acid can be converted into ATP via the normal aerobic respiration pathways

Thyroxin: the Basics A. A hormone produced in the thyroid gland (neck) that controls the metabolic rate (rate of the chem. reactions in the cell) in ALL the cells in your body   B. The more thyroxin present, the greater the metabolic rate C. This will increase sugar and oxygen consumption and also create more body heat